Reduced derivatives of folic acid are required for biosynthesis of DNA, RNA and amino acids. Folic acid is essential for humans, while both plants and microorganisms can synthesize this vitamin de novo. Escherichia coli (E. coli) synthesize folic acid in a pathway that is well characterized. In contrast, the genes or enzymes involved in folate catabolism remain largely unidentified. The abg region of the E. coli chromosome includes three genes that encode proteins that enable uptake and growth on the folate breakdown product, p-aminobenzoyl-glutamate (PABA-GLU). The abg region includes abgA, abgB, abgT, and ogt;these genes may be regulated by AbgR, a divergently-transcribed LysR-type regulator. Prior studies have demonstrated that AbgT catalyzes import of PABA-GLU, while abgA and abgB comprise subunits of PABA-GLU hydrolase. Our broad underlying hypothesis is that this region of the chromosome may be part of a bacterial system for catabolism of the vitamin folic acid. This proposal is designed to further characterize the genes and proteins of this region. Using quantitative real time PCR and Western blot analyses, we propose to investigate the regulation of this putative operon by AbgR. In addition, we will continue to characterize PABA-GLU hydrolase, a novel metal-dependent enzyme. These studies will lead to increased understanding of folate catabolism in E. coli, and may offer alternative targets for development of new antibacterial drugs. In addition, PABA-GLU hydrolase may have clinical applications similar to carboxypeptidase G2, which is used clinically to degrade methotrexate in cancer patients suffering from methotrexate toxicity. PUBLIC HEALTH RELEVANCE: We propose studies on the role of the genes in a little studied area of the E. coli chromosome, the abg region, which is involved in breakdown of the vitamin folic acid. These studies may lead to development of new antibiotics, as well as possibly a new treatment for anti-folate toxicity in chemotherapy patients.